X-ray absorption spectroscopic (XAS) investigations designed to elucidate the electronic and molecular structures of the active sites of a variety of metalloenzymes will be carried out. In each case, structure-function relationships will be developed by determining structural changes accompanying redox and/or substrate- binding steps of the enzyme's catalytic cycle. The ultimate goal is to understand the molecular mechanisms of catalysis by these enzymes. Studies on cytochrome c oxidases from beef heart (cytochrome aa3) and Thermus thermophilous (cytochrome ba3) are designed to elucidate the Cu-Fe binuclear O2-interaction site structure in the resting state and at various stages of turnover (by examining stable analogs of O2 intermediates). Examples of all known types of nickel-containing enzymes, including jackbean urease, Klebsiella aerogenes urease, Desulfovibrio gigas (NiFe) hydrogenase, D. baculatus (NiFeSe) hydrogenase, Clostridium thermoaceticum CO dehydrogenase, and Methanobacterium thermoautotrophicum S-methyl coenzyme-M reductase, will be compared in terms of their nickel site structures. Emphasis will be placed on isolated F430, the nickel-containing tetrapyrrole prosthetic group of methyl reductase, for which ligand-binding studies will help identify any protein-derived axial ligands in methyl reductase. Other physicochemical studies of F430 and methyl reductase will be directed at elucidating the mechanism of CH4 production in this final enzymatic step of methanogenesis. Comparative studies are also planned on amine oxidases from a number of sources to elucidate the local structural environment of the copper sites and their involvement with the organic cofactor in oxidation of primary amines.